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Identification of positions in the human neuropeptide Y/peptide YY receptor Y2 that contribute to pharmacological differences between receptor subtypes
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Pharmacology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Pharmacology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Pharmacology.
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2011 (English)In: Neuropeptides, ISSN 0143-4179, E-ISSN 1532-2785, Vol. 45, no 4, 293-300 p.Article in journal (Refereed) Published
Abstract [en]

The members of the neuropeptide Y (NPY) family are key players in food-intake regulation. In humans this family consists of NPY, peptide YY (PYY) and pancreatic polypeptide (PP) which interact with distinct preference for the four receptors showing very low sequence identity, i.e. Y1, Y2, Y4 and Y5. The binding of similar peptides to these divergent receptors makes them highly interesting for mutagenesis studies. We present here a site-directed mutagenesis study of four amino acid positions in the human Y2 receptor. T(3.40) was selected based on sequence alignments both between subtypes and between species and G(2.68), L(4.60) and Q(6.55) also on previous binding studies of the corresponding positions in the Y1 receptor. The mutated receptors were characterized pharmacologically with the peptide agonists NPY, PYY, PYY(3-36), NPY(13-36) and the non-peptide antagonist BIIE0246. Interestingly, the affinity of NPY and PYY(3-36) increased for the mutants T(3.40)I and Q(6.55)A. Increased affinity was also observed for PYY to Q(6.55)A. PYY(3-36) displayed decreased affinity for G(2.68)N and L(4.60)A whereas binding of NPY(13-36) was unaffected by all mutations. The antagonist BIIE0246 showed decreased affinity for T(3.40)I, L(4.60)A and Q(6.55)A. Although all positions investigated were found important for interaction with at least one of the tested ligands the corresponding positions in hY1 seem to be of greater importance for ligand binding. Furthermore these data indicate that binding of the agonists and the antagonist differs in their points of interaction. The increase in the binding affinity observed may reflect an indirect effect caused by a conformational change of the receptor. These findings will help to improve the structural models of the human NPY receptors.

Place, publisher, year, edition, pages
2011. Vol. 45, no 4, 293-300 p.
Keyword [en]
Y2 receptor, Y1 receptor, Binding experiment, Neuropeptide Y, Peptide YY, BIIE0246, Appetite regulation
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-156633DOI: 10.1016/j.npep.2011.05.006ISI: 000295349600006PubMedID: 21696823OAI: oai:DiVA.org:uu-156633DiVA: diva2:432638
Available from: 2011-08-04 Created: 2011-08-04 Last updated: 2017-12-08Bibliographically approved
In thesis
1. Studies of the Neuropeptide Y Receptor Y2 in Human and Zebrafish
Open this publication in new window or tab >>Studies of the Neuropeptide Y Receptor Y2 in Human and Zebrafish
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The G-protein coupled receptors (GPCRs) comprise the largest family of receptors in humans and other vertebrates. They are embedded in the cell membrane and are activated by many different signaling molecules. Activation modulates cellular signal transduction pathways and influences many physiological processes. Therefore the GPCRs are important as targets for numerous drugs.

The receptors for NPY (neuropeptide Y) belong to GPCRs of Class A (rhodopsin-like). NPY and its related peptides PYY and PP are involved in the regulation of appetite, blood pressure and many other processes. They share a common structure and interact with the receptors Y1, Y2, Y4 and Y5 in mammals, and, in addition, Y7 and Y8 in amphibians and bony fishes.

This thesis is focused on the human Y2 receptor, known to reduce appetite, by investigating the importance of thirteen amino acid residues for ligand binding. Mutagenesis followed by functional expression and receptor binding was conducted. During the course of this work several new GPCR crystal structures have been resolved, thereby improving the receptor modeling in papers I-III. The major finding is that even though the Y1 and Y2 receptors have evolved from a common ancestor, their points of ligand interaction differ and have thus changed during evolution. In general, the positions investigated resulted in milder changes in the ligands’ affinities for Y2 compared to Y1. These findings were incorporated in the design of new Y1 and Y2 receptor models, leading to improved understanding of how such divergent receptors, sharing only 30 percent sequence identity, can still interact with the same ligands. Notably, several of the mutations introduced in Y2 resulted in increased affinity.

A novel NPY receptor gene named Y2-2 was identified in the genomes of zebrafish and medaka. This brings the number of zebrafish NPY receptors to seven. The binding characteristics of zebrafish Y2-2 differed from zebrafish Y2 mainly in the interaction with NPY13-36 and the antagonist BIIE0246.

In conclusion, these results increase our understanding of ligand interactions with GPCRs and will be useful for refinement of ligand-receptor models for future development of receptor subtype-selective drugs.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2011. 55 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 687
Keyword
Y2-receptor, NPY, PYY, G-protein-coupled receptor, mutagenesis, receptor binding, zebrafish, evolution
National Category
Pharmacology and Toxicology
Research subject
Pharmacology
Identifiers
urn:nbn:se:uu:diva-156635 (URN)978-91-554-8117-9 (ISBN)
Public defence
2011-09-21, B:41, Uppsala Biomedical Center (BMC), Husargatan 3, Uppsala, 09:15 (English)
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Available from: 2011-08-31 Created: 2011-08-04 Last updated: 2011-09-08Bibliographically approved

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Gutiérrez-de-Terán, Hugo

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